Bioactive depside and anthocyanin compounds, compositions, and methods of use

ABSTRACT

Methods for modulating the level of a chemokine in a cell by administering to a cell an effective amount of a depside or an anthocyanin are provided. More particularly, a method for modulating the level of a chemokine in a cell by administering to a cell an effective amount of a depside having the structure of formula (IV): Formula (IV) wherein R is selected from H or CH 3  or an anthocyanin selected from cyanidin 3-glucoside, delphinidin 3-glucoside, or combinations thereof, or an enantiomer, optical isomer, diastereomer, N-oxide, crystalline form, hydrate, or pharmaceutically acceptable salt thereof is provided. Also provided are compounds according to Formulas I-IV, pharmaceutical compositions, unit dosage forms, and food or feed supplements containing such compounds. Methods for treating a condition in a mammal and for treating or ameliorating a condition, such as for example, chronic obstructive pulmonary disease (COPD) by administering an effective amount of a composition containing such compounds are also provided. Further provided is an extract obtained from the fruit of  Myrciaria cauliflora  containing at least one compound of the present invention in substantially pure form.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional PatentApplication Ser. No. 60/834,719, filed Jul. 31, 2006. The entiredisclosure of this application is relied upon and incorporated byreference herein.

FIELD OF THE INVENTION

The present invention relates to compounds, compositions, and methodsfor modulating the level of a chemokine in a cell by administering to acell an effective amount of a depside, an anthocyanin, or combinationsthereof. More particularly, the present invention relates to methods forusing the compounds and compositions disclosed herein to treat orameliorate a condition in a mammal, such as for example, chronicobstructive pulmonary disease (COPD).

BACKGROUND OF THE INVENTION

The jaboticaba (Myrciaria cauliflora (Mart.) O. Berg. (Myrtaceae)) is asmall tree native to the Minas Gerais region near Rio de Janeiro insouthern Brazil grown for the purple, grape-like fruits it produces.Traditionally, an astringent decoction of the sun-dried skins has beenused as a treatment for hemoptysis, asthma, diarrhea, and gargled forchronic inflammation of the tonsils (1). The fruit is 3-4 cm in diameterwith one to four large seeds, borne directly on the main trunks andbranches of the plant, lending a distinctive appearance to the fruitingtree. The jaboticaba fruit has a thick, purple, astringent skin thatcovers a sweet, white, gelatinous flesh. Common in Brazilian markets,jaboticabas are largely eaten fresh. Jaboticaba's popularity has beenlikened to that of grapes in the US (2). The fresh fruit may begin toferment 3 to 4 days after harvest, so it is often used to make jams,tarts, strong wines, and liqueurs.

In Brazil, the fruit of several species, namely M. jaboticaba (Veil.) O.Berg, M. tenella (DC.) O. Berg, and M. trunciflora O. Berg, share thesame common name (1-3). The phytochemistry of these fruits has not beenextensively reported in the literature. The jaboticaba (no speciesdistinguished) has been reported to contain tannins (1). The presence ofcyanidin 3-glucoside (Compound 3) in M. cauliflora has been reported(4). M. jaboticaba reportedly contains peonidin 3-glucoside and itsaglycone (5), and the related camu-camu berry (M. dubia), an ediblefruit known for its high levels of ascorbic acid, contains Compound 3and delphinidin 3-glucoside (Compound 4) as its main pigments (6).

Depsides are phenolic compounds composed of two or more monocyclicaromatic units linked by an ester bond. They are most often found inlichens, but have also been isolated from higher plants, includingspecies of the Ericaceae, Lamiaceae, and Papaveraceae (10, 11). Theyhave not been previously reported in the Myrtaceae. Depsides haveantibiotic, anti-HIV, and antiproliferative activity (12, 13). Depsidesalso act as inhibitors of prostaglandin biosynthesis and leukotriene B₄biosynthesis. Depsides are also potent non-steroidal anti-inflammatories(14).

The anthocyanins are a group of well-studied phenolic compounds withantioxidant, anti-inflammatory, antimutagenic, and cancerchemopreventative activities (21). In one study, it was shown thatUVB-exposed HaCaT keratinocytes pretreated with Compound 3 wereprotected from UVB-induced inflammation, inhibiting NF-kB and AP-1activation and IL-8 mRNA expression (22). Depsides from foods andbotanicals are less well-studied than the anthocyanins, possibly as aresult of their limited distribution in higher plants, and their abilityto inhibit IL-8 production and cytotoxcity against colon cancer cellshas not previously been reported.

The jaboticaba is rich in anthocyanins, phenolic acids, flavonoids, andcontains depsides. Accordingly, it would be advantageous to provide,isolate, and characterize the active compound(s) responsible for thestrong antiradical activity of the crude methanolic extracts of thejaboticaba. It would also be advantageous to provide bio-activecompositions containing these bio-active compounds and methods ofadministering them.

SUMMARY OF THE INVENTION

One embodiment of the present invention is method for modulating thelevel of a chemokine in a cell. This method comprises administering to acell an effective amount of a depside, anthocyanin, or a combinationthereof, which is sufficient to modulate the level of a chemokine in acell.

Another embodiment of the present invention is a pharmaceuticalcomposition for treating a condition in a mammal. This method comprisesadministering to the mammal a pharmaceutically acceptable carrier and acompound having the structure of formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof, or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof.

An additional embodiment of the present invention is a method fortreating a condition in a mammal. This method comprises administering tothe mammal an effective amount of a pharmaceutically acceptable carrierand a pharmaceutically acceptable composition comprising a compoundhaving the structure of formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof, or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof.

Another embodiment of the present invention is a method for treating orameliorating chronic obstructive pulmonary disease (COPD). This methodcomprises administering to a mammal an effective amount of a compositioncomprising a pharmaceutically acceptable carrier and a compound havingthe structure of formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof, or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof.

A further embodiment of the present invention is a unit dosage formcomprising a pharmaceutically acceptable carrier and a compound havingthe structure of formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof, or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof.

An additional embodiment of the present invention is a food or feedsupplement comprising an acceptable carrier and a compound havingformula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof, or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof.

Another embodiment of the present invention is an extract obtained fromthe fruit of Myrciaria cauliflora comprising, in substantially pureform, a compound having formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof.

A further embodiment of the present invention is a compound of formula(I):

whereinR₁, R₂, and R₃ are independently selected from H, hydroxy, C₁₋₈alkyl,C₁₋₈alkoxy, C₁₋₈aralkyl, 3- to 8-membered carbocyclic, 3- to 8-memberedheterocyclic, 3- to 8-membered aryl, or 3- to 8-membered heteroaryl,acyl, alkylsulfonyl, and arylsulfonyl, wherein each alkyl, alkoxy,aralkyl, carbocyclic, heterocyclic, aryl, heteroaryl, acyl,alkylsulfonyl, and arylsulfonyl is optionally substituted with at leastone substituent;R₄ is selected from H, C₁₋₈alkyl, C₁₋₈alkoxy, 3- to 8-memberedcarbocyclic, 3- to 8-membered heterocyclic, 3- to 8-membered aryl, or 3-to 8-membered heteroaryl, carboxylate, ester, amide, carbohydrate, aminoacid, acyl, alkoxy-substituted acyl, alditol, NR⁷R⁸, OC(R⁷)₂COOH,SC(R⁷)₂COOH, NHCHR⁷COOH, COR⁸, CO₂R⁸, sulfate, sulfonamide, sulfoxide,sulfonate, sulfone, thioalkyl, thioester, and thioether, wherein eachalkyl, alkoxy, carbocyclic, heterocyclic, aryl, heteroaryl, carboxylate,ester, amide, carbohydrate, amino acid, acyl, alkoxy-substituted acyl,alditol, NR⁷R⁸, OC(R⁷)₂COOH, SC(R⁷)₂COOH, NHCHR⁷COOH, COR⁸, CO₂R⁸,sulfate, sulfonamide, sulfoxide, sulfonate, sulfone, thioalkyl,thioester, and thioether is optionally substituted with at least onesubstituent;R⁷ is selected from H, C₁₋₈alkyl, carbocycle, aryl, heteroaryl,heterocycle, alkylaryl, alkylheteroaryl, and alkylheterocycle, whereineach alkyl, carbocycle, aryl, heteroaryl, heterocycle, alkylaryl,alkylheteroaryl, and alkylheterocycle may be optionally substituted withat least one substituent;R⁸ is selected from H, C₁₋₈alkyl, C₁₋₈alkenyl, C₁₋₈alkynyl, aryl,carbocycle, heteroaryl, heterocycle, alkylaryl, alkylheteroaryl,alkylheterocycle, and heteroaromatic, wherein each alkyl, alkenyl,alkynyl, aryl, carbocycle, heteroaryl, heterocycle, alkylaryl,alkylheteroaryl, alkylheterocycle, and heteroaromatic may be optionallysubstituted with at least one substituent;and n is from 1 to 5,with the proviso that R₁ and R₄ cannot both be H and that when R₁ isOCH₃, R₄ cannot be CH₃; or an enantiomer, optical isomer, diastereomer,N-oxide, crystalline form, hydrate, or pharmaceutically acceptable saltthereof.

An additional embodiment of the present invention is a compound offormula (II):

whereinR₁, R₂, and R₃ are independently selected from H, hydroxy, C₁₋₃alkyl,and C₁₋₃alkoxy, wherein each alkyl and alkoxy is optionally substitutedwith at least one substituent;R₄ is selected from H, C₁₋₃alkyl, and C₁₋₃alkoxy wherein each alkyl andalkoxy, is optionally substituted with at least one substituent;R⁷ is selected from H, carbocycle, aryl, heteroaryl, heterocycle,alkylaryl, alkylheteroaryl, and alkylheterocycle, wherein each alkyl,carbocycle, aryl, heteroaryl, heterocycle, alkylaryl, alkylheteroaryl,and alkylheterocycle may be optionally substituted with at least onesubstituent;R⁸ is selected from H, C₁₋₈alkyl, C₁₋₈alkenyl, C₁₋₈alkynyl, aryl,carbocycle, heteroaryl, heterocycle, alkylaryl, alkylheteroaryl,alkylheterocycle, and heteroaromatic, wherein each alkyl, alkenyl,alkynyl, aryl, carbocycle, heteroaryl, heterocycle, alkylaryl,alkylheteroaryl, alkylheterocycle, and heteroaromatic may be optionallysubstituted with at least one substituent;and n is from 1 to 5,with the proviso that R₁ and R₄ cannot both be H and that when R₁ isOCH₃, R₄ cannot be CH₃; or an enantiomer, optical isomer, diastereomer,N-oxide, crystalline form, hydrate, or pharmaceutically acceptable saltthereof.

Another embodiment of the present invention is a compound of formula(III):

wherein n is 1-5. Preferably, n is 1.

A further embodiment of the present invention is a compound of theformula:

DETAILED DESCRIPTION OF THE INVENTION

The study of antioxidants and cancer chemopreventative compounds fromtropical fruits is ongoing (7-9). Crude methanolic extracts of thejaboticaba have strong antiradical activity in the1,1-diphenyl-2-picrylhydrazyl (DPPH) assay (IC₅₀=35 μg/mL). A newdepside, jaboticabin (Compound 1), was isolated from the crudemethanolic extracts of jaboticaba. In addition, the related depside2-O-(3,4-dihydroxybenzoyl)-2,4,6-trihydroxyphenylacetic acid (Compound2), delphinidin 3-glucoside (Compound 4), pyranocyanin B, quercetin,isoquercitrin, quercimeritrin, quercitrin, rutin, myricitrin, cinnamicacid, O-coumaric acid, gallic acid, protocatechuic acid, methylprotocatechuate, and ellagic acid were identified in this species forthe first time.

The structures of Compounds 1 and 2 are shown below:

wherein R is CH₃ (Compound 1) or H (Compound 2).

Compounds 1 and 2 (depsides), exhibit antiradical activity in the DPPHassay, colon cancer cell cytotoxicity, and significantly inhibitchemokine interleukin (IL)-8 production in human small airway epithelial(SAE) cells before and after treatment with cigarette smoke extract(CSE). Compounds 3 and 4 (anthocyanins), major constituents ofjaboticaba fruits, also display significant activity against IL-8production in SAE cells. Compounds 1-4 were more effective at blockingIL-8 production in untreated SAE cells than catechin. Compounds 2 and 4were more effective than catechin at blocking cigarette smoke-inducedinflammation. Thus, jaboticaba anthocyanins and depsides exhibit goodantiradical activity, cytotoxicity, and inhibit IL-8 production in bothuntreated SAE cells and those treated with proinflammatory CSE. (SeeExamples, below.)

IL-8 is a chemokine implicated in some cancers and a wide range ofchronic inflammatory conditions, including rheumatoid arthritis, andheart and lung diseases (12, 16, 17). The ability of Compounds 1 and 2to reduce IL-8 production suggests that they have an importantanti-inflammatory action.

Chronic obstructive pulmonary disease (COPD) is a complex lung diseasecharacterized by irreversible airflow obstruction due to chronicinflammation. COPD includes chronic obstructive bronchiolitis (fibrosisand obstruction of small airways) and emphysema (permanent enlargementof the airspaces distal to the terminal bronchioles accompanied bydestruction of lung parenchyma). COPD is considered steroid-resistant,and it has been noted that non-steroidal anti-inflammatories that targetchemokine pathways are needed as new therapies (16, 18). The ability ofjaboticaba depsides and anthocyanins, e.g., Compounds 1-4, to reduceinflammation secondary to smoke exposure means these compound havepromise as novel therapies for COPD.

Moreover, the cytotoxicity of Compounds 1, 2, and 4 is comparable toIC₅₀ values for 5-fluorouracil (5-FU), a drug used to treat coloncancer, epigallocatechin gallate (EGCG), and Polyphenon E (Poly E), astandardized decaffeinated green tea extract (8, 19). Thus, thesecompounds show promise as treatments for colon cancer.

Accordingly, one embodiment of the present invention is a method formodulating the level of a chemokine in a cell. This method includesadministering to a cell an effective amount of a depside or anthocyanin,which is sufficient to modulate the level of a chemokine in a cell.

As used herein, the term “depside” means a phenolic compound composed oftwo or more monocyclic aromatic units linked by an ester bond. As usedherein, the term “chemokine” means one of a family of pro-inflammatoryactivation-inducible cytokines, or small protein signals secreted bycells. Chemokines may induce directed chemotaxis in nearby responsivecells, hence the name chemotactic cytokines. Preferably, the chemokineis IL-8.

As used herein the term “anthocyanin” means a water soluble flavonoidpigment that reflects light in the red to blue range of the visiblespectrum, depending on the pH of the surrounding solution. Anthocyaninsare found exclusively in the plant kingdom, and have been observed tooccur in all tissues of higher plants. Preferably, the anthocyanin iscyanidin 3-glucoside, delphinidin 3-glucoside, or combinations thereof,or an enantiomer, optical isomer, diastereomer, N-oxide, crystallineform, hydrate, or pharmaceutically acceptable salt thereof.

As used herein, the term “modulate” means to increase or decrease theamount of, e.g., a chemokine, in a cell as compared to the level of,e.g., the chemokine, in an untreated cell. Preferably, modulate means todecrease the level of the chemokine in the cell.

The present invention also includes compounds of formula (I):

whereinR₁, R₂, and R₃ are independently selected from H, hydroxy, C₁₋₈alkyl,C₁₋₈alkoxy, C₁₋₈aralkyl, 3- to 8-membered carbocyclic, 3- to 8-memberedheterocyclic, 3- to 8-membered aryl, or 3- to 8-membered heteroaryl,acyl, alkylsulfonyl, and arylsulfonyl,wherein each alkyl, alkoxy, aralkyl, carbocyclic, heterocyclic, aryl,heteroaryl, acyl, alkylsulfonyl, and arylsulfonyl is optionallysubstituted with at least one substituent;R₄ is selected from H, C₁₋₈alkyl, C₁₋₈alkoxy, 3- to 8-memberedcarbocyclic, 3- to 8-membered heterocyclic, 3- to 8-membered aryl, or 3-to 8-membered heteroaryl, carboxylate, ester, amide, carbohydrate, aminoacid, acyl, alkoxy-substituted acyl, alditol, NR⁷R⁸, OC(R⁷)₂COOH,SC(R⁷)₂COOH, NHCHR⁷COOH, COR⁸, CO₂R⁸, sulfate, sulfonamide, sulfoxide,sulfonate, sulfone, thioalkyl, thioester, and thioether, wherein eachalkyl, alkoxy, carbocyclic, heterocyclic, aryl, heteroaryl, carboxylate,ester, amide, carbohydrate, amino acid, acyl, alkoxy-substituted acyl,alditol, NR⁷R⁸, OC(R⁷)₂COOH, SC(R⁷)₂COOH, NHCHR⁷COOH, COR⁸, CO₂R⁸,sulfate, sulfonamide, sulfoxide, sulfonate, sulfone, thioalkyl,thioester, and thioether is optionally substituted with at least onesubstituent;R⁷ is selected from H, C₁₋₈alkyl, carbocycle, aryl, heteroaryl,heterocycle, alkylaryl, alkylheteroaryl, and alkylheterocycle, whereineach alkyl, carbocycle, aryl, heteroaryl, heterocycle, alkylaryl,alkylheteroaryl, and alkylheterocycle may be optionally substituted withat least one substituent;R⁸ is selected from H, C₁₋₈alkyl, C₁₋₈alkenyl, C₁₋₈alkynyl, aryl,carbocycle, heteroaryl, heterocycle, alkylaryl, alkylheteroaryl,alkylheterocycle, and heteroaromatic, wherein each alkyl, alkenyl,alkynyl, aryl, carbocycle, heteroaryl, heterocycle, alkylaryl,alkylheteroaryl, alkylheterocycle, and heteroaromatic may be optionallysubstituted with at least one substituent;and n is from 1 to 5,with the proviso that R₁ and R₄ cannot both be H and that when R₁ isOCH₃, R₄ cannot be CH₃; or an enantiomer, optical isomer, diastereomer,N-oxide, crystalline form, hydrate, or pharmaceutically acceptable saltthereof.

In the present invention, each optional substituent is independentlyselected from the group consisting of H, cyano, oxo, nitro, acyl,acylamino, halogen, hydroxy, amino acid, amine, amide, carbamate, ester,ether, C₁₋₈alkanoic acid, carboxylic acid, thio, thioalkyl, thioester,thioether, C₁₋₈alkyl, C₁₋₈alkoxy, C₁₋₈alkenyl, C₁₋₈aralkyl, 3- to8-membered carbocyclic, 3- to 8-membered heterocyclic, 3- to 8-memberedaryl, or 3- to 8-membered heteroaryl, sulfate, sulfonamide, sulfoxide,sulfonate, sulfone, alkylsulfonyl, and arylsulfonyl.

In another embodiment, the present invention includes compounds offormula (II):

whereinR₁, R₂, and R₃ are independently selected from H, hydroxy, C₁₋₃alkyl,and C₁₋₃alkoxy, wherein each alkyl and alkoxy is optionally substitutedwith at least one substituent;R₄ is selected from H, C₁₋₃alkyl, and C₁₋₃alkoxy wherein each alkyl andalkoxy, is optionally substituted with at least one substituent;R⁷ is selected from H, C₁₋₈alkyl, carbocycle, aryl, heteroaryl,heterocycle, alkylaryl, alkylheteroaryl, and alkylheterocycle, whereineach alkyl, carbocycle, aryl, heteroaryl, heterocycle, alkylaryl,alkylheteroaryl, and alkylheterocycle may be optionally substituted withat least one substituent;R⁸ is selected from H, C₁₋₈alkynyl, aryl, carbocycle, heteroaryl,heterocycle, alkylaryl, alkylheteroaryl, alkylheterocycle, andheteroaromatic, wherein each alkyl, alkenyl, alkynyl, aryl, carbocycle,heteroaryl, heterocycle, alkylaryl, alkylheteroaryl, alkylheterocycle,and heteroaromatic may be optionally substituted with at least onesubstituent;and n is from 1 to 5,with the proviso that R₁ and R₄ cannot both be H and that when R₁ isOCH₃, R₄ cannot be CH₃; or an enantiomer, optical isomer, diastereomer,N-oxide, crystalline form, hydrate, or pharmaceutically acceptable saltthereof.

In a further embodiment, the present invention includes compounds offormula (III):

wherein n is 1-5. Preferably, n is 1.

In another embodiment, the present invention includes compounds offormula (IV):

wherein R is selected from H or CH₃ an anthocyanin, combinationsthereof, or an enantiomer, optical isomer, diastereomer, N-oxide,crystalline form, hydrate, or pharmaceutically acceptable salt thereof,which is sufficient to modulate the level of a chemokine in a cell.Preferably, R is CH₃.

As used herein, the term “acyl” has its art-recognized meaning andrefers to a group represented by the general formula hydrocarbylC(O)—,preferably alkylC(O)—.

As used herein, the term “acylamino” has its art-recognized meaning andrefers to an amino group substituted with an acyl group and may berepresented, for example, by the formula hydrocarbylC(O)NH—.

As used herein, the term “acyloxy” has its art-recognized meaning andrefers to a group represented by the general formula hydrocarbylC(O)O—,preferably alkylC(O)O—.

The term “alkoxy” refers to an alkyl group, preferably a lower alkylgroup, having an oxygen attached thereto. Representative alkoxy groupsinclude methoxy, ethoxy, propoxy, tert-butoxy and the like.

The term “alkoxyalkyl” refers to an alkyl group substituted with analkoxy group and may be represented by the general formulaalkyl-O-alkyl.

The term “alkenyl”, as used herein, refers to an aliphatic groupcontaining at least one double bond and is intended to include both“unsubstituted alkenyls” and “substituted alkenyls”, the latter of whichrefers to alkenyl moieties having substituents replacing a hydrogen onone or more carbons of the alkenyl group. Such substituents may occur onone or more carbons that are included or not included in one or moredouble bonds. Moreover, such substituents include all those contemplatedfor alkyl groups, as discussed below, except where stability isprohibitive. For example, substitution of alkenyl groups by one or morealkyl, carbocyclyl, aryl, heterocyclyl, or heteroaryl groups iscontemplated.

The term “alkyl” refers to the radical of saturated aliphatic groups,including straight-chain alkyl groups, branched-chain alkyl groups,cycloalkyl (alicyclic) groups, alkyl-substituted cycloalkyl groups, andcycloalkyl-substituted alkyl groups. In preferred embodiments, astraight chain or branched chain alkyl has 30 or fewer carbon atoms inits backbone (e.g., C₁-C₃₀ for straight chains, C₃-C₃₀ for branchedchains), and more preferably 20 or fewer, such as from 1 to 8. Likewise,preferred cycloalkyls have from 3-10 carbon atoms in their ringstructure, and more preferably have 5, 6 or 7 carbons in the ringstructure.

Moreover, unless otherwise indicated, the term “alkyl” (or “loweralkyl”) as used throughout the specification, examples, and claims isintended to include both “unsubstituted alkyls” and “substitutedalkyls”, the latter of which refers to alkyl moieties havingsubstituents replacing a hydrogen on one or more carbons of thehydrocarbon backbone. Such substituents can include, for example, ahalogen, a hydroxyl, a carbonyl (such as a carboxyl, an alkoxycarbonyl,a formyl, or an acyl), a thiocarbonyl (such as a thioester, athioacetate, or a thioformate), an alkoxyl, a phosphoryl, a phosphate, aphosphonate, a phosphinate, an amino, an amido, an amidine, an imine, acyano, a nitro, an azido, a sulfhydryl, an alkylthio, a sulfate, asulfonate, a sulfamoyl, a sulfonamido, a sulfonyl, a heterocyclyl, anaralkyl, or an aromatic or heteroaromatic moiety. It will be understoodby those skilled in the art that the moieties substituted on thehydrocarbon chain can themselves be substituted, if appropriate. Forinstance, the substituents of a substituted alkyl may includesubstituted and unsubstituted forms of amino, azido, imino, amido,phosphoryl (including phosphonate and phosphinate), sulfonyl (includingsulfate, sulfonamido, sulfamoyl and sulfonate), and silyl groups, aswell as ethers, alkylthios, carbonyls (including ketones, aldehydes,carboxylates, and esters), —CF₃, —CN and the like. Exemplary substitutedalkyls are described below. Cycloalkyls can be further substituted withalkyls, alkenyls, alkoxys, alkylthios, aminoalkyls, carbonyl-substitutedalkyls, —CF₃, —CN, and the like.

The term “C_(x-y)” when used in conjunction with a chemical moiety, suchas, acyl, acyloxy, alkyl, alkenyl, alkynyl, or alkoxy is meant toinclude groups that contain from x to y carbons in the chain. Forexample, the term “C_(x-y)alkyl” refers to substituted or unsubstitutedsaturated hydrocarbon groups, including straight-chain alkyl andbranched-chain alkyl groups that contain from x to y carbons in thechain, including haloalkyl groups such as trifluoromethyl and2,2,2-trifluoroethyl, etc. C₀ alkyl indicates a hydrogen where the groupis in a terminal position, a bond if internal. The terms“C_(2-y)alkenyl” and “C_(2-y)alkynyl” refer to substituted orunsubstituted unsaturated aliphatic groups analogous in length andpossible substitution to the alkyls described above, but that contain atleast one double or triple bond respectively.

The term “alkylthio”, as used herein, refers to a thiol groupsubstituted with an alkyl group and may be represented by the generalformula alkylS—.

The term “alkynyl”, as used herein, refers to an aliphatic groupcontaining at least one triple bond and is intended to include both“unsubstituted alkynyls” and “substituted alkynyls”, the latter of whichrefers to alkynyl moieties having substituents replacing a hydrogen onone or more carbons of the alkynyl group. Such substituents may occur onone or more carbons that are included or not included in one or moretriple bonds. Moreover, such substituents include all those contemplatedfor alkyl groups, as discussed above, except where stability isprohibitive. For example, substitution of alkynyl groups by one or morealkyl, carbocyclyl, aryl, heterocyclyl, or heteroaryl groups iscontemplated.

The term “amide”, as used herein, refers to a group

wherein R⁷ and R⁸ each independently represent a hydrogen or hydrocarbylgroup, or R⁷ and R⁸ taken together with the N atom to which they areattached complete a heterocycle having from 4 to 8 atoms in the ringstructure.

The terms “amine” and “amino” are art-recognized and refer to bothunsubstituted and substituted amines and salts thereof, e.g., a moietythat can be represented by

wherein R⁷, R⁸, and R^(8′) each independently represent a hydrogen or ahydrocarbyl group, or R⁷ and R⁸ taken together with the N atom to whichthey are attached complete a heterocycle having from 4 to 8 atoms in thering structure.

The term “aminoalkyl”, as used herein, refers to an alkyl groupsubstituted with an amino group.

The term “aralkyl”, as used herein, refers to an alkyl group substitutedwith an aryl group.

The term “aryl” as used herein include substituted or unsubstitutedsingle-ring aromatic groups in which each atom of the ring is carbon.Preferably the ring is a 3- to 8-membered ring, more preferably a6-membered ring. The term “aryl” also includes polycyclic ring systemshaving two or more cyclic rings in which two or more carbons are commonto two adjoining rings wherein at least one of the rings is aromatic,e.g., the other cyclic rings can be cycloalkyls, cycloalkenyls,cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls. Aryl groupsinclude benzene, naphthalene, phenanthrene, phenol, aniline, and thelike.

The term “carbamate” is art-recognized and refers to a group

wherein R⁷ and R⁸ independently represent hydrogen or a hydrocarbylgroup.

The terms “carbocycle”, “carbocyclyl”, and “carbocyclic”, as usedherein, refers to a non-aromatic saturated or unsaturated ring in whicheach atom of the ring is carbon. Preferably a carbocycle ring containsfrom 3 to 10 atoms, more preferably from 3 to 8 atoms, including 5 to 7atoms, such as for example, 6 atoms.

The term “carboxy”, as used herein, refers to a group represented by theformula —CO₂H.

The term “ester”, as used herein, refers to a group —C(O)OR⁷ wherein R⁷represents a hydrocarbyl group.

The term “ether”, as used herein, refers to a hydrocarbyl group linkedthrough an oxygen to another hydrocarbyl group. Accordingly, an ethersubstituent of a hydrocarbyl group may be hydrocarbyl-O—. Ethers may beeither symmetrical or unsymmetrical. Examples of ethers include, but arenot limited to, heterocycle-O-heterocycle and aryl-O-heterocycle. Ethersinclude “alkoxyalkyl” groups, which may be represented by the generalformula alkyl-O-alkyl.

The terms “halo” and “halogen” are used interchangeably herein and meanhalogen and include chloro, fluoro, bromo, and iodo.

The terms “hetaralkyl” and “heteroaralkyl”, as used herein, refers to analkyl group substituted with a hetaryl group.

The terms “heteroaryl” and “hetaryl” include substituted orunsubstituted aromatic single ring structures, preferably 3- to8-membered rings, more preferably 5- to 7-membered rings, even morepreferably 5- to 6-membered rings, whose ring structures include atleast one heteroatom, preferably one to four heteroatoms, morepreferably one or two heteroatoms. The terms “heteroaryl” and “hetaryl”also include polycyclic ring systems having two or more cyclic rings inwhich two or more carbons are common to two adjoining rings wherein atleast one of the rings is heteroaromatic, e.g., the other cyclic ringscan be cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls,and/or heterocyclyls. Heteroaryl groups include, for example, pyrrole,furan, thiophene, imidazole, oxazole, thiazole, pyrazole, pyridine,pyrazine, pyridazine, and pyrimidine, and the like.

The term “heteroatom” as used herein means an atom of any element otherthan carbon or hydrogen. Preferred heteroatoms are nitrogen, oxygen, andsulfur.

The terms “heterocyclyl”, “heterocycle”, and “heterocyclic” refer tosubstituted or unsubstituted non-aromatic ring structures, preferably 3-to 10-membered rings, more preferably 3- to 8-membered rings, whose ringstructures include at least one heteroatom, preferably one to fourheteroatoms, more preferably one or two heteroatoms. The terms“heterocyclyl” and “heterocyclic” also include polycyclic ring systemshaving two or more cyclic rings in which two or more carbons are commonto two adjoining rings wherein at least one of the rings isheterocyclic, e.g., the other cyclic rings can be cycloalkyls,cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls.Heterocyclyl groups include, for example, piperidine, piperazine,pyrrolidine, morpholine, lactones, lactams, and the like.

The term “heterocyclylalky”, as used herein, refers to an alkyl groupsubstituted with a heterocycle group.

The term “hydrocarbyl”, as used herein, refers to a group that is bondedthrough a carbon atom that does not have a ═O or ═S substituent, andtypically has at least one carbon-hydrogen bond and a primarily carbonbackbone, but may optionally include heteroatoms. Thus, groups likemethyl, ethoxyethyl, 2-pyridyl, and trifluoromethyl are considered to behydrocarbyl for the purposes of this application, but substituents suchas acetyl (which has a ═O substituent on the linking carbon) and ethoxy(which is linked through oxygen, not carbon) are not. Hydrocarbyl groupsinclude, but are not limited to aryl, heteroaryl, carbocycle,heterocycle, alkyl, alkenyl, alkynyl, and combinations thereof.

The term “hydroxyalkyl”, as used herein, refers to an alkyl groupsubstituted with a hydroxy group.

The term lower when used in conjunction with a chemical moiety, such as,acyl, acyloxy, alkyl, alkenyl, alkynyl, or alkoxy is meant to includegroups where there are ten or fewer non-hydrogen atoms in thesubstituent, preferably eight or fewer, such as for example, from about3 to 8 carbon atoms, more preferably less than 6 carbon atoms. A “loweralkyl”, for example, refers to an alkyl group that contains ten or fewercarbon atoms, preferably six or fewer. In certain embodiments, acyl,acyloxy, alkyl, alkenyl, alkynyl, or alkoxy substituents defined hereinare respectively lower acyl, lower acyloxy, lower alkyl, lower alkenyl,lower alkynyl, or lower alkoxy, whether they appear alone or incombination with other substituents, such as in the recitationshydroxyalkyl and aralkyl (in which case, for example, the atoms withinthe aryl group are not counted when counting the carbon atoms in thealkyl substituent).

The terms “polycyclyl”, “polycycle”, and “polycyclic” refer to two ormore rings (e.g., cycloalkyls, cycloalkenyls, cycloalkynyls, aryls,heteroaryls, and/or heterocyclyls) in which two or more atoms are commonto two adjoining rings, e.g., the rings are “fused rings”. Each of therings of the polycycle can be substituted or unsubstituted. In certainembodiments, each ring of the polycycle contains from 3 to 10 atoms inthe ring, preferably from 3 to 8, such as for example, 5 to 7.

The term “substituted” refers to moieties having substituents replacinga hydrogen on one or more carbons of the backbone. It will be understoodthat “substitution” or “substituted with” includes the implicit provisothat such substitution is in accordance with permitted valence of thesubstituted atom and the substituent, and that the substitution resultsin a stable compound, e.g., which does not spontaneously undergotransformation such as by rearrangement, cyclization, elimination, etc.As used herein, the term “substituted” is contemplated to include allpermissible substituents of organic compounds. In a broad aspect, thepermissible substituents include acyclic and cyclic, branched andunbranched, carbocyclic and heterocyclic, aromatic and non-aromaticsubstituents of organic compounds. The permissible substituents can beone or more and the same or different for appropriate organic compounds.For purposes of this invention, the heteroatoms such as nitrogen mayhave hydrogen substituents and/or any permissible substituents oforganic compounds described herein which satisfy the valences of theheteroatoms. Substituents can include any substituents described herein,for example, a halogen, a hydroxyl, a carbonyl (such as a carboxyl, analkoxycarbonyl, a formyl, or an acyl), a thiocarbonyl (such as athioester, a thioacetate, or a thioformate), an alkoxyl, a phosphoryl, aphosphate, a phosphonate, a phosphinate, an amino, an amido, an amidine,an imine, a cyano, a nitro, an azido, a sulfhydryl, an alkylthio, asulfate, a sulfonate, a sulfamoyl, a sulfonamido, a sulfonyl, aheterocyclyl, an aralkyl, or an aromatic or heteroaromatic moiety. Itwill be understood by those skilled in the art that the moietiessubstituted on the hydrocarbon chain can themselves be substituted, ifappropriate.

Unless specifically stated as “unsubstituted,” references to chemicalmoieties herein are understood to include substituted variants. Forexample, reference to an “aryl” group or moiety implicitly includes bothsubstituted and unsubstituted variants.

The term “sulfate” is art-recognized and refers to the group —OSO₃H, ora pharmaceutically acceptable salt thereof.

The term “sulfonamide” is art-recognized and refers to the grouprepresented by the general formulae

wherein R⁷ and R⁸ independently represents hydrogen or hydrocarbyl.

The term “sulfoxide” is art-recognized and refers to the group —S(O)—R⁷,wherein R⁷ represents a hydrocarbyl.

The term “sulfonate” is art-recognized and refers to the group SO₃H, ora pharmaceutically acceptable salt thereof.

The term “sulfone” is art-recognized and refers to the group —S(O)₂—R⁷,wherein R⁷ represents a hydrocarbyl.

The term “thioalkyl”, as used herein, refers to an alkyl groupsubstituted with a thiol group.

The term “thioester”, as used herein, refers to a group —C(O)SR⁷ or—SC(O)R⁷ wherein R⁷ represents a hydrocarbyl.

The term “thioether”, as used herein, is equivalent to an ether, whereinthe oxygen is replaced with a sulfur.

In the present invention, the compounds of the present invention may beisolated from natural sources, such as, e.g., in the case of Compounds 1and 2 from the jaboticaba. Alternatively, Compound 2 may be isolatedfrom, e.g., the dried petals of P rhoeas L. See, e.g., Hillenbrand M. etal., Planta Med, 70:378-380 (2004). Alternatively, any of the compoundswithin the scope of formulae I-III may be synthesized using routineskill in the art. See, e.g., Kuisle O. et al., J Org Chem.,64(22):8063-8075 (1999) and Kuisle O. et al., Tetrahedron Letters,40(6):1203-1206 (1999).

The present invention also includes compositions, includingpharmaceutical compositions, containing any of the foregoing compounds.The present invention further includes methods of treating orameliorating a condition as defined below, particularly COPD, byadministering to a mammal an effective amount of any of the foregoingcompounds or compositions.

A further embodiment of the present invention is a pharmaceuticalcomposition for treating a condition in a mammal. In this embodiment,the composition contains a pharmaceutically acceptable carrier and acompound having the structure of formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof, or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof. Preferably, R isCH₃.

An additional embodiment of the present invention is a method fortreating a condition in a mammal. This method includes administering tothe mammal a pharmaceutically acceptable carrier and an effective amountof a pharmaceutically acceptable composition comprising a compoundhaving the structure of formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof, or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof, and apharmaceutically acceptable carrier. Preferably, R is CH₃.

As used herein, the term “condition” means a symptomatic state or adisease in the mammal. Preferably, the condition is cancer, aninflammatory condition, or both. Preferably, the inflammatory conditionis a chronic inflammatory condition. Preferably, the chronicinflammatory condition is rheumatoid arthritis, heart disease, or lungdisease. Preferably, the lung disease is COPD. Preferably, the cancer iscolon cancer.

In the present invention, an “effective amount” is an amount sufficientto effect beneficial or desired results. An effective amount can beadministered in one or more doses. In terms of treatment, an “effectiveamount” of a depside or anthocyanin for a cell is an amount sufficientto modulate the level of a chemokine in a cell.

In terms of treatment of a mammal, an “effective amount” of thecompounds of the present invention (i.e., Compounds 1-4) is an amountsufficient to treat, manage, palliate, ameliorate, or stabilize acondition in the mammal. More particularly, an “effective amount”delivers to the subject from about 1 to about 1000 mg, preferably fromabout 5 to about 500 mg, more preferably from about 10 to about 250 mg,such as for example, about 20, 40, 60, 80, 100, 120, 140, 160, 180, 200,220, or 240 mg of one of the compounds of the present invention.

The effective amount is generally determined by a physician on acase-by-case basis and is within the skill of one in the art. Severalfactors are typically taken into account when determining an appropriatedosage. These factors include age, sex and weight of the patient, thecondition being treated, the severity of the condition and the form ofthe drug being administered. For instance, the amount of a derivative ofa compound of formula (I) may not need to be as high as that of thecompound of formula (I) itself in order to be therapeutically effective.

Effective dosage forms, modes of administration, and dosage amounts maybe determined empirically, and making such determinations is within theskill of the art. It is understood by those skilled in the art that thedosage amount will vary with the route of administration, the rate ofexcretion, the duration of the treatment, the identity of any otherdrugs being administered, the age, size, and species of animal, and likefactors well known in the arts of medicine and veterinary medicine. Ingeneral, a suitable dose of one of the compounds according to theinvention will be that amount of the compound, which is the lowest doseeffective to produce the desired effect. The effective dose of acompound according to the invention maybe administered as two, three,four, five, six or more sub-doses, administered separately atappropriate intervals throughout the day.

A compound of the present invention may be administered in any desiredand effective manner: as pharmaceutical compositions for oral ingestion,or for parenteral or other administration in any appropriate manner suchas intraperitoneal, subcutaneous, topical, intradermal, inhalation,intrapulmonary, rectal, vaginal, sublingual, intramuscular, intravenous,intraarterial, intrathecal, or intralymphatic. Further, a compound ofthe invention may be administered in any combination with each otherand/or in conjunction with other treatments. A compound of inventionmaybe encapsulated or otherwise protected against gastric or othersecretions, if desired.

In an embodiment of the present invention, the compound or compositionof the present invention is formulated for aerosol administration,particularly to the respiratory tract. The compound or compositiongenerally has a small particle size, e.g., about 5 microns or less. Sucha particle size may be obtained by means known in the art, for exampleby micronization. The particles of the compound or composition areprovided in a pressurized pack with a suitable propellant such as achlorofluorocarbon (CFC), for example dichlorodifluoromethane,trichlorofluoromethane, or dichlorotetrafluoroethane, carbon dioxide, orother suitable gas. The aerosol may conveniently also contain asurfactant such as lecithin. The dose of drug may be controlled by ametered valve. Alternatively, the compound or composition may beprovided in a form of a dry powder, for example a powder mix of thecompound in a suitable powder base such as lactose, starch, starchderivatives such as hydroxypropylmethyl cellulose andpolyvinylpyrrolidine (PVP). The powder composition may be presented inunit dose form, for example in capsules or cartridges of, e.g., gelatinor blister packs from which the powder may be administered by means ofan inhaler. See, e.g., Berg, Published International Application No. WO2002/009699; Sedwell, R. W. et al., Chemptherapy, 40(1):43-56 (1994).

While it is possible for a compound of the invention to be administeredalone, it is preferable to administer the compound(s) as apharmaceutical formulation (composition). The pharmaceuticallyacceptable compositions comprise one or more compounds of the presentinvention as an active ingredient in admixture with one or morepharmaceutically-acceptable carriers and, optionally, one or more othercompounds, drugs, ingredients and/or materials. Regardless of the routeof administration selected, the compounds of the present invention areformulated into pharmaceutically-acceptable dosage forms by conventionalmethods known to those of skill in the art. See, e.g., Remington'sPharmaceutical Sciences (Mack Publishing Co., Easton, Pa.).

Another aspect of this embodiment is a method for killing a cancer cell.In this method, an effective amount of the pharmaceutical composition asdefined above is administered. Preferably, the cancer cell is a coloncancer cell. In another aspect of this method, the pharmaceuticalcomposition is conjointly administered with at least one agent.

In the present invention, one or more compounds of the present inventionmay be conjointly administered to the mammal with an agent that killscells through, e.g., an apoptotic mechanism. In this aspect, the agentis, e.g., a chemotherapeutic agent—including combinations ofchemotherapeutic agents. Agents useful in the present invention include,e.g., an EGF-receptor antagonist, arsenic sulfide, adriamycin,cisplatin, carboplatin, cimetidine, carminomycin, mechlorethaminehydrochloride, pentamethylmelamine, thiotepa, teniposide,cyclophosphamide, chlorambucil, demethoxyhypocrellin A, melphalan,ifosfamide, trofosfamide, Treosulfan, podophyllotoxin orpodophyllotoxin, derivatives, etoposide phosphate, teniposide,etoposide, leurosidine, leurosine, vindesine, 9-aminocamptothecin,camptoirinotecan, crisnatol, megestrol, methopterin, mitomycin C,ecteinascidin 743, busulfan, carmustine (BCNU), lomustine (CCNU),lovastatin, 1-methyl-4-phenylpyridinium ion, semustine, staurosporine,streptozocin, phthalocyanine, dacarbazine, aminopterin, methotrexate,trimetrexate, thioguanine, mercaptopurine, fludarabine, pentastatin,cladribin, cytarabine (ara C), porfiromycin, 5-fluorouracil,6-mercaptopurine, doxorubicin hydrochloride, leucovorin, mycophenolicacid, daunorubicin, deferoxamine, floxuridine, doxifluridine,raltitrexed, idarubicin, epirubican, pirarubican, zorubicin,mitoxantrone, bleomycin sulfate, actinomycin D, safracins, saframycins,quinocarcins, discodermolides, vincristine, vinblastine, vinorelbinetartrate, vertoporfin, paclitaxel, tamoxifen, raloxifene, tiazofuran,thioguanine, ribavirin, EICAR, estramustine, estramustine phosphatesodium, flutamide, bicalutamide, buserelin, leuprolide, pteridines,enediynes, levamisole, aflacon, interferon, interleukins, aldesleukin,filgrastim, sargramostim, rituximab, BCG, tretinoin, betamethosone,gemcitabine hydrochloride, verapamil, VP-16, altretamine, thapsigargin,oxaliplatin, iproplatin, tetraplatin, lobaplatin, DCP, PLD-147, JM118,JM216, JM335, satraplatin, docetaxel, deoxygenated paclitaxel, TL-139,5′-nor-anhydrovinblastine (hereinafter: 5′-nor-vinblastine),camptothecin, irinotecan (Camptosar, CPT-11), topotecan (Hycamptin), BAY38-3441, 9-nitrocamptothecin (Orethecin, rubitecan), exatecan (DX-8951),lurtotecan (GI-147211C), gimatecan, homocamptothecins diflomotecan(BN-80915) and 9-aminocamptothecin (IDEC-13′), SN-38, ST1481,karanitecin (BNP1350), indolocarbazoles (e.g., NB-506), protoberberines,intoplicines, idenoisoquinolones, benzo-phenazines or NB-506.

Pharmaceutical carriers are well known in the art (see, e.g.,Remington's Pharmaceutical Sciences (Mack Publishing Co., Easton, Pa.)and The National Formulary (American Pharmaceutical Association,Washington, D.C.)) and include sugars (e.g., lactose, sucrose, mannitol,and sorbitol), starches, cellulose preparations, calcium phosphates(e.g., dicalcium phosphate, tricalcium phosphate and calcium hydrogenphosphate), sodium citrate, water, aqueous solutions (e.g., saline,sodium chloride injection, Ringer's injection, dextrose injection,dextrose and sodium chloride injection, lactated Ringers injection),alcohols (e.g., ethyl alcohol, propyl alcohol, and benzyl alcohol),polyols (e.g., glycerol, propylene glycol, and polyethylene glycol),organic esters (e.g., ethyl oleate and tryglycerides), biodegradablepolymers (e.g., polylactide-polyglycolide, poly(orthoesters), andpoly(anhydrides)), elastomeric matrices, liposomes, microspheres, oils(e.g., corn, germ, olive, castor, sesame, cottonseed, and groundnut),cocoa butter, waxes (e.g., suppository waxes), paraffins, silicones,talc, silicylate, etc. Each carrier used in a pharmaceutical compositionof the invention must be “acceptable” in the sense of being compatiblewith the other ingredients of the formulation and not injurious to thesubject. Carriers suitable for a selected dosage form and intended routeof administration are well known in the art, and acceptable carriers fora chosen dosage form and method of administration can be determinedusing ordinary skill in the art.

The pharmaceutically acceptable compositions (or acceptable compositionsin the case of, e.g., feed supplements) of the invention may,optionally, contain additional ingredients and/or materials commonlyused in pharmaceutical compositions. These ingredients and materials arewell known in the art and include (1) fillers or extenders, such asstarches, lactose, sucrose, glucose, mannitol, and silicic acid; (2)binders, such as carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, hydroxypropylmethyl cellulose, sucrose and acacia; (3)humectants, such as glycerol; (4) disintegrating agents, such asagar-agar, calcium carbonate, potato or tapioca starch, alginic acid,certain silicates, sodium starch glycolate, cross-linked sodiumcarboxymethyl cellulose and sodium carbonate; (5) solution retardingagents, such as paraffin; (6) absorption accelerators, such asquaternary ammonium compounds; (7) wetting agents, such as cetyl alcoholand glycerol monosterate; (8) absorbents, such as kaolin and bentoniteclay; (9) lubricants, such as talc, calcium stearate, magnesiumstearate, solid polyethylene glycols, and sodium lauryl sulfate; (10)suspending agents, such as ethoxylated isostearyl alcohols,polyoxyethylene sorbitol and sorbitan esters, microcrystallinecellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth;(11) buffering agents; (12) excipients, such as lactose, milk sugars,polyethylene glycols, animal and vegetable fats, oils, waxes, paraffins,cocoa butter, starches, tragacanth, cellulose derivatives, polyethyleneglycol, silicones, bentonites, silicic acid, talc, salicylate, zincoxide, aluminum hydroxide, calcium silicates, and polyamide powder; (13)inert diluents, such as water or other solvents; (14) preservatives;(15) surface-active agents; (16) dispersing agents; (17) control-releaseor absorption-delaying agents, such as hydroxypropylmethyl cellulose,other polymer matrices, biodegradable polymers, liposomes, microspheres,aluminum monosterate, gelatin, and waxes; (18) opacifying agents; (19)adjuvants; (20) wetting agents; (21) emulsifying and suspending agents;(22), solubilizing agents and emulsifiers, such as ethyl alcohol,isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol,benzyl benzoate, propylene glycol, 1,3-butylene glycol, oils (inparticular, cottonseed, groundnut, corn, germ, olive, castor and sesameoils), glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fattyacid esters of sorbitan; (23) propellants, such aschlorofluorohydrocarbons and volatile unsubstituted hydrocarbons, suchas butane and propane; (24) antioxidants; (25) agents which render theformulation isotonic with the blood of the intended recipient, such assugars and sodium chloride; (26) thickening agents; (27) coatingmaterials, such as lecithin; and (28) sweetening, flavoring, coloring,perfuming and preservative agents. Each such ingredient or material mustbe “acceptable” in the sense of being compatible with the otheringredients of the formulation and not injurious to the subject.Ingredients and materials suitable for a selected dosage form andintended route of administration are well known in the art, andacceptable ingredients and materials for a chosen dosage form and methodof administration may be determined using ordinary skill in the art.

Pharmaceutical formulations suitable for oral administration may be inthe form of capsules, cachets, pills, tablets, powders, granules, asolution or a suspension in an aqueous or non-aqueous liquid, anoil-in-water or water-in-oil liquid emulsion, an elixir or syrup, apastille, a bolus, an electuary or a paste. These formulations may beprepared by methods known in the art, e.g., by means of conventionalpan-coating, mixing, granulation or lyophilization processes.

Solid dosage forms for oral administration (capsules, tablets, pills,dragees, powders, granules and the like) may be prepared by mixing theactive ingredient(s) with one or more pharmaceutically-acceptablecarriers and, optionally, one or more fillers, extenders, binders,humectants, disintegrating agents, solution retarding agents, absorptionaccelerators, wetting agents, absorbents, lubricants, and/or coloringagents. Solid compositions of a similar type maybe employed as fillersin soft and hard-filled gelatin capsules using a suitable excipient. Atablet may be made by compression or molding, optionally with one ormore accessory ingredients. Compressed tablets may be prepared using asuitable binder, lubricant, inert diluent, preservative, disintegrant,surface-active or dispersing agent. Molded tablets may be made bymolding in a suitable machine. The tablets, and other solid dosageforms, such as dragees, capsules, pills and granules, may optionally bescored or prepared with coatings and shells, such as enteric coatingsand other coatings well known in the pharmaceutical-formulating art.They may also be formulated so as to provide slow or controlled releaseof the active ingredient therein. These compositions may also optionallycontain opacifying agents and may be of a composition such that theyrelease the active ingredient only, or preferentially, in a certainportion of the gastrointestinal tract, optionally, in a delayed manner.The active ingredient can also be in microencapsulated form.

Liquid dosage forms for oral administration includepharmaceutically-acceptable emulsions, microemulsions, solutions,suspensions, syrups and elixirs. The liquid dosage forms may containsuitable inert diluents commonly used in the art. Besides inertdiluents, the oral compositions may also include adjuvants, such aswetting agents, emulsifying and suspending agents, sweetening,flavoring, coloring, perfuming and preservative agents. Suspensions maycontain suspending agents.

Formulations for rectal or vaginal administration may be presented as asuppository, which maybe prepared by mixing one or more activeingredient(s) with one or more suitable nonirritating carriers which aresolid at room temperature, but liquid at body temperature and,therefore, will melt in the rectum or vaginal cavity and release theactive compound. Formulations which are suitable for vaginaladministration also include pessaries, tampons, creams, gels, pastes,foams or spray formulations containing such pharmaceutically-acceptablecarriers as are known in the art to be appropriate.

Dosage forms for the topical or transdermal administration includepowders, sprays, ointments, pastes, creams, lotions, gels, solutions,patches, drops and inhalants. The active compound may be mixed understerile conditions with a suitable pharmaceutically-acceptable carrier.The ointments, pastes, creams and gels may contain excipients. Powdersand sprays may contain excipients and propellants.

Pharmaceutical compositions suitable for parenteral administrationscomprise one or more compounds of the present invention in combinationwith one or more pharmaceutically-acceptable sterile isotonic aqueous ornon-aqueous solutions, dispersions, suspensions or emulsions, or sterilepowders which may be reconstituted into sterile injectable solutions ordispersions just prior to use, which may contain suitable antioxidants,buffers, solutes which render the formulation isotonic with the blood ofthe intended recipient, or suspending or thickening agents. Properfluidity can be maintained, for example, by the use of coatingmaterials, by the maintenance of the required particle size in the caseof dispersions, and by the use of surfactants. These compositions mayalso contain suitable adjuvants, such as wetting agents, emulsifyingagents and dispersing agents. It may also be desirable to includeisotonic agents. In addition, prolonged absorption of the injectablepharmaceutical form may be brought about by the inclusion of agentswhich delay absorption.

In some cases, in order to prolong the effect of a drug, it is desirableto slow its absorption from subcutaneous or intramuscular injection.This may be accomplished by the use of a liquid suspension ofcrystalline or amorphous material having poor water solubility.

The rate of absorption of the drug then depends upon its rate ofdissolution which, in turn, may depend upon crystal size and crystallineform. Alternatively, delayed absorption of a parenterally-administereddrug may be accomplished by dissolving or suspending the drug in an oilvehicle. Injectable depot forms may be made by forming microencapsulematrices of the active ingredient in biodegradable polymers. Dependingon the ratio of the active ingredient to polymer, and the nature of theparticular polymer employed, the rate of active ingredient release canbe controlled. Depot injectable formulations are also prepared byentrapping the drug in liposomes or microemulsions which are compatiblewith body tissue. The injectable materials can be sterilized forexample, by filtration through a bacterial-retaining filter.

The formulations may be presented in unit-dose or multi-dose sealedcontainers, for example, ampules and vials, and may be stored in alyophilized condition requiring only the addition of the sterile liquidcarrier, for example water for injection, immediately prior to use.Extemporaneous injection solutions and suspensions may be prepared fromsterile powders, granules and tablets of the type described above.

Another embodiment of the present invention is a method for treating orameliorating chronic obstructive pulmonary disease (COPD). This methodincludes administering to a mammal an effective amount of a compositioncomprising a pharmaceutically acceptable carrier and a compound havingthe structure of formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof, or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof. In this method,the mammal is preferably a human. It is also preferred that R is CH₃.

A further embodiment of the present invention is a unit dosage formcomprising a pharmaceutically acceptable carrier and a compound havingthe structure of formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof, or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof. In this method, itis preferred that R is CH₃.

An additional embodiment of the present invention is a food or feedsupplement comprising an acceptable carrier, such as for example, apharmaceutically acceptable carrier as defined above, and a compoundhaving formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof, or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof. In this method, itis preferred that R is CH₃.

Another embodiment of the present invention is an extract obtained fromthe fruit of Myrciaria cauliflora comprising, in substantially pureform, a compound having formula (IV):

wherein R is selected from H or CH₃ or a compound selected from cyanidin3-glucoside, delphinidin 3-glucoside, or combinations thereof.Preferably, R is CH₃.

In the present invention, “substantially pure form” means that theextract contains at least one compound of the present invention that isat least 60% pure, more suitably at least 75% pure and preferably atleast 85%, especially at least 98% pure (% are on a weight for weightbasis). Impure preparations of the compounds may be used for preparingthe more pure forms used in the pharmaceutical compositions.

An aspect of this embodiment is a food or feed supplement (as describedabove) containing the extract. Another aspect of this embodiment is apharmaceutical composition containing the extract and a pharmaceuticallyacceptable carrier, as described above.

Another aspect of this embodiment is a method for treating orameliorating a condition in a mammal comprising administering to amammal an effective amount of a composition containing the extract.Preferably, the mammal is a human.

The conditions treated, forms and formulations of the composition,methods of administration and delivery routes for the composition, andeffective doses are as defined above.

The following examples are provided to further illustrate thecompositions and methods of the present invention. These examples areillustrative only and are not intended to limit the scope of theinvention in any way.

Examples I. General Experimental Procedures

UV spectra were measured on a Perkin-Elmer Lambda 35 UV/VISspectrometer. NMR experiments were conducted on a Bruker Avance AV300NMR spectrometer operating at 300.13 MHz for ¹H and 75.48 MHz for ¹³Cusing standard Bruker software. Mass spectra were obtained on aThermoFinnigan LCQ utilizing both ESI and APCI in the positive andnegative modes. HRESIMS was performed on a Micromass Q-TOF Ultima massspectrometer. HPLC was done on a Waters 2695 using a Phenomenex Aquacolumn (250×4.6 mm, 5 μm) and monitored using a Waters 996 PDA scanningfrom 240 to 600 nm. Column chromatography was accomplished usingSephadex LH-20 (Pharmacia, 25-100 μm), reversed-phase C18 silica gel (J.T. Baker, 40 μm), and Diaion HP-20 (Mitsubishi, Japan). Separations weremonitored using silica gel 60 F254 and RP18 F254 TLC plates (1 mmthickness, EM Science, Germany). Quercetin, rutin, cinnamic acid,O-coumaric acid, gallic acid, and 1,1-diphenyl-2-picrylhydrazyl (DPPH)were purchased from Sigma (MO, USA). Isoquercitrin and myricitrin werepreviously isolated in the laboratory (23).

II. Plant Material

Fruits of M. cauliflora were collected at the Fruit and Spice Park inHomestead, Fla., immediately frozen and shipped by overnight courier ondry ice to the laboratory, where they were kept in cold (−20° C.), darkstorage until processed. A voucher specimen (Reynertson 39) wasprepared, identified, and deposited at the Steere Herbarium of The NewYork Botanical Garden (Bronx, N.Y.).

III. Extraction and Isolation Procedures

Deseeded fresh fruits (6.2 kg) were homogenized in a blender with MeOH,extracted exhaustively and concentrated in vacuo at temperatures notexceeding 40° C. to give a thick syrup that was diluted with water. Theaqueous solution was separated over Diaion HP-20 and eluted using H₂O,MeOH, and acetone. The MeOH fraction was concentrated to give a residue(52 g). A portion (44 g) of that residue was subjected to Sephadex LH-20column chromatography (about 125 g) in amounts of 8, 11, 12, and 13 gand eluted with formic acid:water:MeOH (1:9:10). Fractions from all fourcolumns were recombined to give 8 fractions (A-H). Fractions D (3.74 g)and E (111 mg) were chromatographed over Sephadex LH-20 and eluted usingMeOH:formic acid (9:1). The recombined fraction A1 was then separated ina smaller Sephadex LH-20 column (12 g) using an isocratic system ofacetonitrile-water. Fractions A1₃₅₋₅₉ were recombined as fraction A2 andsubjected to reversed-phase C18 column chromatography (6 g) using 10%formic acid:acetonitrile (95:5 to 50:50; 5% gradient; 20 ml each eluant,fractions of 3 mL). Fractions A2₁₂₋₂₇ were recombined and subjected tofinal purification using Sephadex LH-20 and water-acetonitrile to give16 mg of Compound 1. Compound 2 was isolated according to an analogousscheme to give 10 mg.

Approximately 50 g of freeze-dried fruits were extracted in EtOH andsubjected to LC-MS selected ion monitoring (SIM) analysis to determineif Compound 1 was a methyl ester artifact of Compound 2 followingextraction in MeOH. Analysis was performed in negative ESI mode, using agradient of 0.1% formic acid (A) and acetonitrile (B) from 95% A to 50%A over 30 minutes, monitoring [M-H]⁻ m/z 332 to 334. A [M-H]⁻ molecularion m/z=333 with the same retention time as Compound 1 was detected inethanolic extracts, indicating that Compound 1 is produced by the plantitself.

A. Jaboticabin (methyl2-[(3,4-dihydroxybenzoyloxy)-4,6-dihydroxyphenyl]acetate)

From reference: Compound 1 is a reddish amorphous solid (MeOH); UV(MeOH) λ_(max) (log ε) 267.5 (3.35), 298.5 (3.16) nm; ¹H NMR (CD₃OD,300.13 MHz) δ 7.55 (1H, dd, J=2.1, 7.8 Hz, H-6′), 7.54 (1H, d, J=2.1 Hz,H-2′), 6.88 (1H, d, J=7.8 Hz, H-5′), 6.27 (1H, d, J=2.4 Hz, H-5), 6.16(1H, d, J=2.4 Hz, H-3), 3.57 (3H, s, OCH₃-8), 3.50 (2H, s, H-7); ¹³C NMR(CD₃OD, 75.48 MHz) 5172.9 (C, C-8), 164.9 (C, C-7′), 166.4 (C-2′), 157.2(C, C-4), 157.0 (C, C-2), 151.1 (C, C-6), 145.0 (C, C-3′), 122.9 (CH,C-6′), 120.3 (C, C-1′), 114.7 (CH, C-5′), 105.7 (C, C-1), 100.7 (CH,C-5), 99.6 (CH, C-3), 50.9 (OCH₃-8), 28.5 (CH₂, C-7); ESIMS m/z 333[M-H]⁻ (C₁₆H₁₄O₈), HRESIMS m/z 357.0581 [M+Na]⁺ (calculated forC₁₆H₁₄O₈Na, 357.0586).

Compound 1 was isolated as a reddish amorphous powder. The negative ESImass spectrum showed a [M-H]⁻ molecular ion of m/z=333. Positive HRESIMSgave a [M+Na]⁺ molecular ion of m/z=357.0581, corresponding to amolecular formula of C₁₆H₁₄O₈. The UV spectrum exhibited a peak at 267nm with a shoulder at 298 nm, typical of a phenolic acid ester. The ¹H-and ¹³C-NMR experiments were similar to literature values for Compound 2with the addition of a methoxy signal at δ 3.57 (3H, s, OCH₃-8) and 50.9(OCH₃-8) (11). The position of the methoxy group was established throughHMBC correlations between the proton signal at δ 3.57 (OCH₃-8) and δ172.9 (C-8). The C-1 attachment for the methyl acetate group followedfrom HMBC correlations; the methylene proton signal at δ 3.50 (H-7)showed HMBC correlations with C-1, C-2, C-6, and the carbonyl C-8. Adetailed analysis of 1-D and 2-D NMR spectra and comparison withCompound 2 confirmed the structure of jaboticabin (Compound 1) as methyl2-[(3,4-dihydroxybenzoyloxy)-4,6-dihydroxyphenyl]acetate:

An ethanolic extract of jaboticaba fruits was analyzed by LC-MS in SIMmode to determine if Compound 1 was a methyl ester artifact from theinitial MeOH extraction. A [M-H]⁻ molecular ion m/z=333 with the sameretention time as Compound 1 was detected in ethanolic extracts, furtherconfirming that Compound 1 is produced by the plant itself.

B. Other Isolated Compounds

Compounds 2, 3, 4, pyranocyanin B, protocatechuic acid, methylprotocatechuate, ellagic acid, quercimeritrin, and quercitrin wereisolated and identified by comparison of spectroscopic measurements topublished literature values (11, 15). Quercetin, isoquercitrin, rutin,myricitrin, cinnamic acid, 0-coumaric acid, and gallic acid wereidentified by comparison of retention time, UV, and MS data to authenticstandards.

IV. 1,1-Diphenyl-2-picrylhydrazyl (DPPH) Assay

The DPPH assay was performed on extracts, fractions, and purifiedcompounds as previously described, using 400 μM DPPH (9). Gallic acidwas used as a positive control (IC₅₀=30.0±2.9 μM). The results arereported in Table 1:

TABLE 1 DPPH assay results. DPPH Assay Compound IC₅₀ (μM) Compound 151.4 Compound 2 61.8 Compound 3 28.4 Compound 4 26.3 Gallic acid(Control) 30.0

The DPPH assay is an indication of the ability of a substance toscavenge free radicals. Gallic acid is know to be an effective freeradical scavenger. Accordingly, the results in Table 1 demonstrate thatCompounds 1-4 exhibit good antiradical activity.

V. IL-8 Immunoassay

Human SAE cells were cultured according to supplier instructions(Clonetics, CA) and maintained in a controlled atmosphere of air-5% CO₂at 37° C. Confluent SAE cells at passages 4-8 were used for experiments.

Cigarette smoke extract (CSE) was prepared using a modified protocol(24). Briefly, a Barnet vacuum pump operating at constant flow was usedto draw the smoke of one unfiltered 2R1 reference cigarette (Universityof Kentucky) through 25 mL of Dulbecco's phosphate-buffered saline. Thissolution (100% CSE) was adjusted to pH 7.4, filtered, diluted with smallairway growth medium to a final concentration of 5%, and added to thecells immediately.

Cells were treated with 5% CSE or pure compounds (100 μM), or pretreatedwith pure compounds 30 minutes prior to 5% CSE exposure. After 24 hoursmeasurement of human IL-8 in cell culture supernates was performed byELISA (R&D Systems Inc., MN). Statistical analyses were performed byStudent's t-test (two-sided) using the JMP Statistics software package(SAS Institute Inc., NC) and defined at the 5% level. The results arereported in Table 2:

TABLE 2 IL-8 immunoassay results. IL-8 inhibition in SAE cells CompoundUntreated Treated with 5% CSE Compound 1 81.3% 47.3% Compound 2 74.9%70.3% Compound 3 65.3% 36.4% Compound 4 Not Detected 96.0% Catechin NoChange 60.3%

As shown in Table 2, Compound 1 decreases IL-8 production in untreatedSAE cells by 81.3% and decreases IL-8 production in SAE cells treatedwith 5% CSE by 47.3%. Compound 2 inhibits IL-8 production by 74.9% inuntreated SAE cells and 70.3% in treated SAE cells.

IL-8 is not detected in SAE cells treated with Compound 4. Compound 4causes a 96% reduction in IL-8 production in SAE cells treated with CSE.Compound 3 inhibits IL-8 production by 65.3% (untreated) and 36.4%(treated), respectively.

VI. Cytotoxicity Assays

Colon cancer cell lines HT29, HCT116, and SW480 (10,000 cells) wereplated into 24-well plates in Dulbecco's modified Eagle medium (DMEM)containing 10% fetal bovine serum (FBS). After 24 hours, cells weretreated with 6 concentrations (1, 5, 10, 30, 50, and 70 μM) of Compounds1-4 and 5-fluorouracil (5-FU) and incubated for 72 hours under DMEMcontaining 1% FBS. The plates were washed with PBS once and the attachedcells were collected by tripsinization. The numbers of cells werecounted using a Coulter Counter (Beckman Coulter Co., CA) as previouslydescribed (25). The results are reported in Table 3:

TABLE 3 Cytotoxicity assays results. IC₅₀ (μM) Compound HT29 HCT116SW480 Compound 1 65 >100 not tested Compound 2 >100 30 not testedCompound 3 ~100 ~70 ~90-100 Compound 4 ~100 12 20 5-FU 46.1 45.1 53.0

The cytotoxicity of Compounds 1, 2, and 4 is comparable to IC₅₀ valuesfor 5-FU, a drug used for colon cancer treatment and to the positivecontrols epigallocatechin gallate (EGCG) and Polyphenon E (Poly E), astandardized decaffeinated green tea extract (8, 19). EGCG and Poly Eexhibited HT29 IC₅₀ of 27 μg/mL and 22 μg/mL, respectively. Compound 1is cytotoxic against HT29 colon cancer cells (IC₅₀=65 μM). Compound 2 iscytotoxic against HCT116 colon cancer cells (IC₅₀=30 μM). Consistentwith published literature, Compound 4 was more cytotoxic than Compound 3(20). Compound 4 showed good activity against both the HCT116 and SW480cell lines (IC₅₀=12 and 20 μM, respectively), while Compound 3 inhibited50% cell growth only at the 100 μM range.

CITED DOCUMENTS

The following documents, cited above, are incorporated by reference asif recited in full herein:

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The scope of the present invention is not limited by the description,examples, and suggested uses herein and modifications can be madewithout departing from the spirit of the invention. Thus, it is intendedthat the present invention cover modifications and variations of thisinvention provided that they come within the scope of the appendedclaims and their equivalents.

1.-39. (canceled)
 40. A compound of formula (I):

wherein R₁, R₂, and R₃ are independently selected from H, hydroxy,C₁₋₈alkoxy, C₁₋₈aralkyl, 3- to 8-membered carbocyclic, 3- to 8-memberedheterocyclic, 3- to 8-membered aryl, or 3- to 8-membered heteroaryl,acyl, alkylsulfonyl, and arylsulfonyl, wherein each alkyl, alkoxy,aralkyl, carbocyclic, heterocyclic, aryl, heteroaryl, acyl,alkylsulfonyl, and arylsulfonyl is optionally substituted with at leastone substituent; R₄ is selected from H, C₁₋₈alkyl, C₁₋₈alkoxy, 3- to8-membered carbocyclic, 3- to 8-membered heterocyclic, 3- to 8-memberedaryl, or 3- to 8-membered heteroaryl, carboxylate, ester, amide,carbohydrate, amino acid, acyl, alkoxy-substituted acyl, alditol, NR⁷R⁸,OC(R⁷)₂COOH, SC(R⁷)₂COOH, NHCHR⁷COOH, COR⁸, CO₂R⁸, sulfate, sulfonamide,sulfoxide, sulfonate, sulfone, thioalkyl, thioester, and thioether,wherein each alkyl, alkoxy, carbocyclic, heterocyclic, aryl, heteroaryl,carboxylate, ester, amide, carbohydrate, amino acid, acyl,alkoxy-substituted acyl, alditol, NR⁷R⁸, OC(R⁷)₂COOH, SC(R⁷)₂COOH,NHCHR⁷COOH, COR⁸, CO₂R⁸, sulfate, sulfonamide, sulfoxide, sulfonate,sulfone, thioalkyl, thioester, and thioether is optionally substitutedwith at least one substituent; R⁷ is selected from H, C₁₋₈alkyl,carbocycle, aryl, heteroaryl, heterocycle, alkylaryl, alkylheteroaryl,and alkylheterocycle, wherein each alkyl, carbocycle, aryl, heteroaryl,heterocycle, alkylaryl, alkylheteroaryl, and alkylheterocycle may beoptionally substituted with at least one substituent; R⁸ is selectedfrom H, C₁₋₈alkenyl, C₁₋₈alkynyl, aryl, carbocycle, heteroaryl,heterocycle, alkylaryl, alkylheteroaryl, alkylheterocycle, andheteroaromatic, wherein each alkyl, alkenyl, alkynyl, aryl, carbocycle,heteroaryl, heterocycle, alkylaryl, alkylheteroaryl, alkylheterocycle,and heteroaromatic may be optionally substituted with at least onesubstituent; and n is from 1 to 5, with the proviso that R₁ and R₄cannot both be H and that when R₁ is OCH₃, R₄ cannot be CH₃; or anenantiomer, optical isomer, diastereomer, N-oxide, crystalline form,hydrate, or pharmaceutically acceptable salt thereof.
 41. Apharmaceutical composition comprising a compound according to claim 40and a pharmaceutically acceptable carrier.
 42. (canceled)
 43. A compoundof formula (II):

wherein R₁, R₂, and R₃ are independently selected from H, hydroxy,C₁₋₃alkyl, and C₁₋₃alkoxy, wherein each alkyl and alkoxy is optionallysubstituted with at least one substituent; R₄ is selected from H,C₁₋₃alkyl, and C₁₋₃alkoxy wherein each alkyl and alkoxy, is optionallysubstituted with at least one substituent; R⁷ is selected from H,C₁₋₈alkyl, carbocycle, aryl, heteroaryl, heterocycle, alkylaryl,alkylheteroaryl, and alkylheterocycle, wherein each alkyl, carbocycle,aryl, heteroaryl, heterocycle, alkylaryl, alkylheteroaryl, andalkylheterocycle may be optionally substituted with at least onesubstituent; R⁸ is selected from H, C₁₋₈alkenyl, C₁₋₈alkynyl, aryl,carbocycle, heteroaryl, heterocycle, alkylaryl, alkylheteroaryl,alkylheterocycle, and heteroaromatic, wherein each alkyl, alkenyl,alkynyl, aryl, carbocycle, heteroaryl, heterocycle, alkylaryl,alkylheteroaryl, alkylheterocycle, and heteroaromatic may be optionallysubstituted with at least one substituent; and n is from 1 to 5, withthe proviso that R₁ and R₄ cannot both be H and that when R₁ is OCH₃, R₄cannot be CH₃; or an enantiomer, optical isomer, diastereomer, N-oxide,crystalline form, hydrate, or pharmaceutically acceptable salt thereof.44. A pharmaceutical composition comprising a compound according toclaim 43 and a pharmaceutically acceptable carrier.
 45. (canceled)
 46. Acompound of formula (III):

wherein n is 1-5.
 47. A pharmaceutical composition comprising a compoundaccording to claim 46 and a pharmaceutically acceptable carrier. 48.(canceled)
 49. A compound of the formula: